Heat shock proteins (HSPs) are found in virtually all prokaryotic and eukaryotic cells where they support folding of nascent polypeptides, prevent protein aggregation, and assist transport of other proteins across membranes. The proteins in the Hsp70 family (referred to collectively as “Hsp70”) play a dual role of protecting cells from lethal damage after environmental stress, on the one hand, and targeting cells for immune mediated cytolytic attack on the other hand. Increased expression of Hsp70 in the cytoplasma is known to protect a broad range of cells under stress by preventing the misfolding, aggregation and denaturation of cytoplasmic proteins and inhibiting various apoptotic pathways. Mosser, et al., Mol Cell Biol., 2000 October; 20(19): 7146-7159; Yenari, Adv. Exp. Med. Biol., 2002, 513, 281-299; Kiang & Tsokos, Pharmacol Ther., 1998; 80(2):182-201. However, membrane-bound Hsp70 provides a target structure for cytolytic attack mediated by natural killer cells.
Cells can experience stress due to temperature; injury (trauma); genetic disease; metabolic defects; apoptosis; infection; toxins; radiation; oxidants; excess/lack of nutrients or metabolic products; and the like. For example, it is known in the art that cells damaged in the following variety of medical conditions can experience a protective effect in response to Hsp70.
Protein misfolding/aggregation conditions resulting in neurodegeneration include Alzheimers' disease (Zhang, et al., J. Neuroscience, 2004, 24(23), 5315-5321; Klettner, Drug News Perspect, 2004, 17(5), 299-306); Huntington's disease (Klettner, ibid); Parkinson's disease (Auluck, et al., Science, 2002, 295(5556), 865-868); and the like. Other neurodegenerative conditions include spinal/bulbar muscular atrophy (Sobue, Nihon Shinkei Seishin Yakurigaku Zasshi, 2001, 21(1), 21-25); and familial amyotrophic lateral sclerosis (Howland, et al., Proc Nat Acad Sci USA, 2002, 99(3), 1604-1609; Sobue, ibid; Vleminck, et al., J. Neuropathol. Exp. Neurol., 2002, 61(11), 968-974).
Ischemia and associated oxidative damage affects diverse tissues including: neurons and glia (Carmel, et al., Exp. Neurol., 2004, 185(1) 81-96; Renshaw & Warburton, Front Biosci., 2004, 9, 110-116; Yenari, Adv. Exp. Med. Biol., 2002, 513, 281-299; Kelly & Yenari, Curr. Res. Med. Opin., 2002, 18 Sup. 2, s55-60; Lee, et al., Exp. Neurol., 2001, 170(1), 129-139; Klettner, ibid; Klettner & Herdegen, Br. J. Pharmacol., 2003, 138(5), 1004-1012); cardiac muscle (Marber, M. S., et al., J. Clin. Invest., (1995) 95:1446-1456; Plumier, J. C., et al., J. Clin. Invest., (1995) 95:1854-1860; Radford, N. B., et al., Proc. Natl. Acad. Sci. USA, (1996), 93(6): 2339-2342; Voss, et al., Am. J. Physiol. Heart Circ. Physiol., 2003, 285: H687-H692); liver tissue (Doi, et al., Hepatogastroenterology, 2001 March-April; 48(38):533-40; Gao, et al., World J. Gastroenterol., 2004; 10(7):1019-1027); skeletal muscle (Lepore et al., Cell Stress & Chaperones, 2001, 6(2), 93-96); kidney tissue (Chen, et al., Kidney Int., 1999; 56: 1270-1273; Beck, et al., Am. J. Physiol. Renal Physiol., 2000, 279: F203-F215); pulmonary tissue (Hiratsuka, et al., J. Heart Lung Transplant, 1998 December; 17(12):1238-46); pancreatic tissue (Bellmann, et al., J. Clin. Invest., 1995 June; 95(6): 2840-2845), and the like.
Seizure conditions that damage neurons include, e.g., epileptic seizure (Yenari, ibid; Blondeau, et al., Neuroscience, 2002, 109(2), 231-241); or chemically induced seizure (Tsuchiya, et al., Neurosurgery, 2003, 53(5), 1179-1187).
Thermal stresses include hyperthermia conditions such as fever, heat stroke, and the like (Barclay & Robertson, J. Neurobiol., 2003 56(4), 360-271; Sato, et al., Brain Res., 1996, 740(1-2), 117-123); and hypothermia (Kandor & Goldberg, Proc. Natl. Acad. Sci. USA, 1997 May 13; 94(10): 4978-4981).
Aging includes conditions such as atherosclerosis which affects smooth muscle cells (Minowada, G. & Welch, W. J., J. Clin. Invest., (1995), 95:3-12; Johnson, A. J., et al., Arterio. Thromb. Vasc. Biol., (1995), 15(1):27-36).
Other conditions include radiation damage, e.g., from ultraviolet light to tissues such as murine fibroblasts (Simon, M. M., et al., J. Clin. Res., (1995), 95(3): 926-933), and light damage to retinal cells (Yu, et al, Molecular Vision, 2001; 7:48-56).
Trauma includes, for example, mechanical injury, e.g., pressure damage to retinal ganglions in glaucoma (Ishii, et al., Invest. Opthalmol. Vis. Sci., 2003, 44(5), 1982-1992).
Toxic conditions include doses of chemicals or biochemicals, for example, methamphetamine (Malberg & Seiden, Poster “MDMA Administration Induces Expression of HSP70 in the Rat Brain” Society for Neuroscience Annual Meeting, New Orleans, La., Oct. 25-30, 1997); antiretroviral HIV therapeutics (Keswani, et al., Annals Neurology, 2002, 53(1), 57-64); heavy metals, amino acid analogs, chemical oxidants, ethanol, glutamate, and other toxins (Ashburner, M. & Bonner, J. J., Cell, (1979) 17:241-254; Lindquist, S., Ann. Rev. Biochem., (1986) 55:1151-1191; Craig, E. A., Crit. Rev. Biochem. (1985), 18(3):239-280; MORIMOTO, et al., THE BIOLOGY OF HEAT SHOCK PROTEINS AND MOLECULAR CHAPERONE, (Cold Spring Harbor Laboratory Press 1994), 417-455); and the like.
Cystic fibrosis is a genetic disorder which results from a mutation in a single glycoprotein called the cystic fibrosis transmembrane conductance regulator (CFTR). As a result of the mutation, post-translational processing of CFTR cannot proceed correctly and the glycoprotein fails to be delivered to the cell membrane. Induction of Hsp70 has been shown to overcome this defective processing and results in functional CFTR protein on the cell surface (Choo-Kang & Zeitlin, Am. J. Physiol. Lung Cell Mol. Physiol., (2001), 281:L58-L68).
Therefore, there is a need for new methods of increasing expression of Hsp70 in order to treat disorders responsive to Hsp70.
Extracellular Hsp70 and membrane bound Hsp70 have been shown to play key roles in activation of the innate immune system. Monocytes have been shown to secrete proinflammatory cytokines in response to soluble Hsp70 protein and membrane bound Hsp70 has been shown to provide a target structure for cytolytic attack by natural killer cell.
Natural killer (NK) cells, a type of white blood cell, are known to be an important component of the body's immune system. Because the defining function of NK cells is spontaneous cytotoxicity without prior immunization, NK cells can be the first line of defense in the immune system, and are believed to play a role in attacking cancer cells and infectious diseases. Many conditions, such as immunodeficiency diseases, aging, toxin exposure, endometriosis, and the like can leave subjects with lowered NK cell activity or dysfunctional NK cells.
For example, subjects can have decreased or deficient NK cell activity, in conditions such as chronic fatigue syndrome (chronic fatigue immune dysfunction syndrome) or Epstein-Barr virus, post viral fatigue syndrome, post-transplantation syndrome or host-graft disease, exposure to drugs such as anticancer agents or nitric oxide synthase inhibitors, natural aging, and various immunodeficiency conditions such as severe combined immunodeficiency, variable immunodeficiency syndrome, and the like. (Caligiuri, et al., J. of Immunology, 1987; 139: 3306-13; Morrison, et al., Clinical and Experimental Immunology, 1991; 83: 441-6; Klingemann, Biology of Blood and Marrow Transplantation, 2000, 6:90-99; Ruggeri, et al., Best Pract Res Clin Haematol., 2004, 17(3):427-38; Cifone, et al., Int. Immunopharmacol., 2001, 1(8):1513-24; Plackett, et al., J. Leukoc. Biol., 2004 August, 76(2):291-9; Alpdogan & van den Brink, Trends Immunol., 2005 January, 26(1):56-64; Heusel & Ballas Z K, Curr. Opin. Pediatr., 2003 December, 15(6):586-93; Hacein-Bey-Abina, et al., Int. J. Hematol., 2002 November, 76(4):295-8; Baumert, et al., Immun Infekt., 1992 July, 20(3):73-5).
NK cells are known to have activity against a wide range of infectious pathogens such as bacteria, viruses, fungi, protozoan parasites, combined infections, e.g., combined bacterial/viral infections, and the like. NK cells are believed to be particularly important in combating intracellular infections where the pathogens replicate in the subjects cells, e.g., a substantial fraction of viruses and many other pathogens that can form intracellular infections.
For example, a wide range of fungal infections are reported to be targeted by NK cells such as Cryptococcus neoformans, dermatophytes, e.g., Trichophyton rubrum, Candida albicans, Coccidioides immitis, Paracoccidioides brasiliensis, or the like (Hidore, et al., Infect. Immun., 1991 April, 59(4):1489-99; Akiba, et al., Eur. J. Dermatol., 2001 January-February, 11(1):58-62; Mathews & Witek-Janusek, J. Med. Microbiol., 1998 November, 47(11):1007-14; Ampel, et al., Infect. Immun., 1992 October, 60(10):4200-4; Jimenez & Murphy, Infect. Immun., 1984 November, 46(2):552-8.)
Also targeted by NK cells are bacteria, especially intracellular bacteria, e.g., Mycobacterium tuberculosis, Mycobacterium avium, Listeria monocytogenes, many different viruses, such as human immunodeficiency virus, herpesviruses, hepatitis, and the like, and viral/bacterial co-infection (Esin et al., Clin. Exp. Immunol., 1996 June, 104(3):419-25; Kaufmann, Annu. Rev. Immunol., 1993, 11:129-63; See et al., Scand. J. Immunol., 1997 September, 46(3):217-24; Brenner et al., J. Leukoc. Biol., 1989 July, 46(1):75-83; Kottilil S., Indian J. Exp. Biol., 2003 November, 41(11):1219-25; Herman & Koziel, Clin. Gastroenterol. Hepatol., 2004 December, 2(12):1061-3; Beadling & Slifka, Curr. Opin. Infect. Dis., 2004 June, 17(3):185-91).
In addition, NK cells combat protozoal infections including toxoplasmosis, trypanosomiasis, leishmaniasis and malaria, especially intracellular infections (Korbel, et al., Int. J. Parasitol., 2004 December, 34(13-14):1517-28; Ahmed & Mehlhorn, Parasitol. Res., 1999 July, 85(7):539-49; Osman, et al., Dig. Surg., 1998, 15(4):287-96; Gazzinelli, et al., Infect. Agents Dis., 1993 June, 2(3):139-49; Askonas & Bancroft, Philos. Trans. R. Soc. Lond. B: Biol. Sci., 1984 Nov. 13, 307(1131):41-9; Allison & Eugui, Annu. Rev. Immunol., 1983; 1:361-92).
NK cells have been shown to play a role in attacking cancer cells that present membrane bound Hsp70. It is believed that membrane bound Hsp70 binds to CD94 receptors on the surface of NK cells to cause them to produce and secrete high amounts of the enzyme granzyme B which is thought to enter the tumor cell via interaction with membrane bound Hsp70 and induce apoptosis (Radons & Multhoff, Exerc. Immunol. Rev., (2005), 11:17-33). Therefore, there is an urgent need for effective treatments for increasing NK cell activity for the treatment of cancer and other disorders that respond to NK induction.